KR850006547A - Method for preparing bovine growth hormone derivative expression vector - Google Patents

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Description

Method for preparing bovine growth hormone derivative expression vector

As this is a public information case, the full text was not included.

1 to 3 are schematic diagrams of the production protocol of plasmid PNM 575.

Claims (34)

The following genes (1), (2), (3), (4), (5), (6), (7), or () which encode runaway replicons and bioactive bovine growth hormones A method of making a selectable and spontaneously replicating DAN expression vector characterized by linking transcriptional and translational activation sequences present in the readout of 8). Wherein A is deoxyadenyl, G is deoxyguanyl, C is deoxycytosyl, T is thymidyl, and R is amino acid 9 (leucine) to 191 (phenylalanine) of bovine growth hormone. DNA sequence to encode, and R ¹ is a DNA sequence encoding the following translation stop sequence. The method of claim 1, wherein the transcriptional activation sequence is E. coli. E. coli lactose, bacteriophage λ P _L O _L , bacteriophage λP _R O _R , tac. this. E. coli lipoprotein or E. coli lipoprotein. E. coli tryptophan transcriptional activation sequence. The method of claim 1 or 2, wherein the vector is a plasmid. The method of claim 3, wherein the translational activating sequence is an E. coli lipoprotein sequence. The method of claim 4, wherein the plasmid PCZ 1920 is linked to a ˜10.2 kb BamHI-XbaI fragment of plasmid PCZ101, a DNA linker having the following nucleotide sequence, and a ˜0.6 kb BamHI-XbaI fragment of plasmid PCZ101. The method of claim 4, wherein the plasmid pJRI is prepared by linking a ˜10.2 kb BamHI-XbaI fragment of plasmid PCZ101, a DNA linker having the following nucleotide sequence, and a ˜0.6 kb BamHI-XbaI fragment of plasmid PCZ101. 5. The method of claim 4, wherein the plasmid pAT ₂ is linked to a ~ 10.2 kb BamHI-XbaI fragment of plasmid pCZ101, a DNA linker of the following nucleotide sequence, and a ˜0.6 kb BamHI-XbaI fragment of plasmid pCZ101. The method of claim 4, wherein the plasmid PJR 1.3 is linked to a 9.3 kb BamHI-XbaI fragment of plasmid pCZ103, a DNA linker of the following nucleotide sequence and a ˜0.6 kb BamHI-XbaI fragment of plasmid pCZ101. The method of claim 3, wherein the translational activation sequence is E. coli. The coli tryptophan sequence. The method of claim 9, wherein the plasmid pCZ112 is linked to a ˜10.2 kb BamHI-EcoRI fragment of plasmid pCZ101, a DNA linker having the following nucleotide sequence, and an EcoRI-claI digest of plasmid NM 608. 11. The method of claim 9, for preparing the plasmid pAT _1, characterized by a connection Sikkim the EcoRI-ClaI digested product of the 10.2kb BamHI-EcoRI fragment of plasmid pCZ101 ~, to the nucleotide sequence of plasmid DNA linker and the NM 608. The method of claim 9, wherein the plasmid pASP ₁ is prepared by linking a ˜10.2 kb BamHI-EcoRI fragment of plasmid pCZ101, a DNA linker having the following nucleotide sequence, and an EcoRI-ClaI digest of plasmid NM 608. 11. The method of claim 9, for preparing the plasmid pASP _2, characterized by a connection Sikkim the EcoRI-ClaI digested product of the 10.2kb BamHI-EcoRI fragment of plasmid pCZ1001 ~, to the nucleotide sequence of plasmid DNA linker and the NM 608. 10. The method of claim 9, wherein the plasmid pCZ154 is linked to a ˜10.2 kb BamHI-EcoRI fragment of plasmid pCZ101, a DNA linker of the following nucleotide sequence, and an EcoRI-ClaI digest of plasmid NM 608. 11. 10. The method of claim 9, wherein the plasmid pCZ155 is linked to a ˜10.2 kb BamHI-EcoRI fragment of plasmid pCZ101, a DNA linker of the following nucleotide sequence, and an EcoRI-ClaI digest of plasmid NM 608. 11. 10. The method of claim 9, wherein the plasmid pCZ156 is linked to a ˜10.2 kb BamHI-EcoRI fragment of plasmid pCEZ101, a DNA linker of the following nucleotide sequence, and an EcoRI-ClaI digest of plasmid NM 608. 11. The method of claim 9, wherein the plasmid pAT2.3 is prepared by linking a 9.3 cd BamHI-EcoRI fragment of plasmid pCZ103, a DNA linker of the following nucleotide sequence and a BamHI-XbaI digest of ˜0.6 kb of plasmid pCZ101. . 10. The method of claim 9, wherein the 9.3 kb BamHI-EcoRI fragment of plasmid pCZ103, the DNA linker of the following nucleotide sequence, and the EcoRI-ClaI digest of plasmid NM608 are linked. The method of claim 9, wherein the plasmid pASP1.3 is prepared by linking a ~ 9.3 kb BamHI-EcoRI fragment of plasmid pCZ103, a DNA linker of the following nucleotide sequence, and an EcoRI-ClaI digest of plasmid NM608. The method of claim 9, wherein the plasmid pASP2.3 is prepared by linking a ~ 9.3 kb BamHI-EcoRI fragment of plasmid pCZ103, a DNA linker of the following nucleotide sequence, and an EcoRI-ClaI digest of plasmid NM608. 10. The method of claim 9, wherein the 9.3 kb BamHI-EcoRI fragment of plasmid pCEZ103, the DNA linker of the following nucleotide sequence and the EcoRI-ClaI digest of plasmid NM608 are linked. 10. The method of claim 9, wherein the 9.3 kb BamHI-EcoRI fragment of plasmid pCZ103, the DNA linker of the following nucleotide sequence and the EcoRI-ClaI digest of plasmid NM 608 are linked. 10. The method of claim 9, wherein the 9.3 kb BamHI-EcoRI fragment of plasmid pCZ103, the DNA linker of the following nucleotide sequence and the EcoRI-ClaI digest of plasmid NM608 are linked. Plasmid pCZ101 is treated with BstEII restriction enzyme, the resulting DNA is purified, and then the purified DNA is linked to generate plasmid pCZ103, which is suitable for use in the method of any one of claims 8 and 17 and 23. How to prepare. present in the readout of a nucleotide sequence encoding a functional polypeptide containing a) a runaway replicon and b) a translational stop signal underlying the nucleotide triplet encoding the carboxy terminus of the functional polypeptide. Transforming the campant cells with a selectable and spontaneously replicable recombinant DNA expression vector consisting of transcriptional and translational activation sequences, and then propagating the transformed cells appropriate for gene expression and activation or induction of runaway replicons. A method for producing protein granules having high intracellular homogeneity, characterized by culturing under conditions. The method of claim 25, wherein the recombinant DNA expression vector is a plasmid. 27. The method of claim 25 or 26, wherein the transcriptional activation sequence is E. coli. E. coli tryptophan transcriptional activation sequence, E. E. coli lipoprotein transcriptional activation sequence, E. E. coli lactose transcriptional activation sequence, bacteriophage λP _L O _L transcriptional activation sequence, or bacteriophage λP _R O _R transcriptional activation sequence. The method of claim 25, 26 or 27, wherein the transcriptional activation sequence is one or more of E. coli. A method of containing E. coli transcriptional activation sequence in series. The method of claim 27, wherein the transcriptional activation sequence is E. coli. E. coli tryptophan transcriptional activation sequence or E. coli. E. coli lipoprotein transcriptional activation sequence. 30. The method according to any one of claims 25 to 29, wherein the functional polypeptide encoding sequence is bovine growth hormone, human growth hormone, human pre-growth hormone, pig growth hormone, mammalian growth hormone, avian growth hormone, human insulin A chain. , Human insulin B chain, human proinsulin, human prepro insulin, interferon, urokinase, human tissue plasminogen activator, growth hormone free factor or sequence encoding interleukin II. The method of any one of claims 26 to 30, wherein the plasmid is plasmid pCZ1920, pJR1, pCZ112, pAT1, pAT2, pASP1, pASP2, pCZ01, pJR1.3, pAT1.3, pAT2.3, pA2SP1.3, pASP2.3, pCZ154 .3, pCZ155.3 or pCZ156.3. 32. The method of any one of claims 25-31, wherein the transformed host cell is a prokaryotic cell. 33. The method of claim 32, wherein the transformed host cell is E. coli. Coli way. The method of claim 33, wherein the transformed host cell is E. coli. E. coli K12 RV308. ※ Note: The disclosure is based on the initial application.